`First published November 13, 2013; doi:10.1152/ajprenal.00525.2013.
`
`Review
`
`Tuberous sclerosis complex, mTOR, and the kidney: report of an
`NIDDK-sponsored workshop
`
`Elizabeth P. Henske,1 Rebekah Rasooly,2 Brian Siroky,3 and John Bissler4
`1Division of Pulmonary and Critical Care Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston,
`Massachusetts; 2Division of Kidney, Urologic, and Hematologic Diseases, National Institute of Diabetes and Digestive and
`Kidney Diseases, National Institutes of Health, Bethesda, Maryland; 3Division of Nephrology and Hypertension, Cincinnati
`Children’s Hospital Medical Center, University of Cincinnati, Cincinnati, Ohio; and 4Tuberous Sclerosis Complex Center of
`Excellence, Le Bonheur Children’s Hospital, University of Tennessee College of Medicine, Memphis Tennessee
`
`Submitted 9 October 2013; accepted in final form 7 November 2013
`
`Downloaded from
`
`http://ajprenal.physiology.org/
`
` by 10.220.33.4 on January 8, 2017
`
`Henske EP, Rasooly R, Siroky B, Bissler J. Tuberous sclerosis complex,
`mTOR, and the kidney: report of an NIDDK-sponsored workshop. Am J Physiol
`Renal Physiol 306: F279 –F283, 2014. First published November 13, 2013;
`doi:10.1152/ajprenal.00525.2013.—Remarkable basic and translational advances
`have elucidated the role of the mammalian target of rapamycin (mTOR) signaling
`network in the pathogenesis of renal disease. Many of these advances originated
`from studies of the genetic disease tuberous sclerosis complex (TSC), leading to
`one of the clearest therapeutic opportunities to target mTOR with rapamycin and its
`analogs (“rapalogs”), which effectively inhibit mTOR complex 1 (mTORC1) by an
`allosteric mechanism. Clinical trials based on these discoveries have provided
`strongly positive therapeutic results in TSC (Bissler JJ, McCormack FX, Young
`LR, Elwing JM, Chuck G, Leonard JM, Schmithorst VJ, Laor T, Brody AS, Bean
`J, Salisbury S, Franz DN. N Engl J Med 358: 140 –151, 2008; Krueger DA, Care
`MM, Holland K, Agricola K, Tudor C, Mangeshkar P, Wilson KA, Byars A,
`Sahmoud T, Franz DN. N Engl J Med 363: 1801–1811, 2010; McCormack FX,
`Inoue Y, Moss J, Singer LG, Strange C, Nakata K, Barker AF, Chapman JT,
`Brantly ML, Stocks JM, Brown KK, Lynch JP 3rd, Goldberg HJ, Young LR,
`Kinder BW, Downey GP, Sullivan EJ, Colby TV, McKay RT, Cohen MM, Korbee
`L, Taveira-DaSilva AM, Lee HS, Krischer JP, Trapnell BC. N Engl J Med 364:
`1595–1606, 2011). In June 2013, the National Institute of Diabetes and Digestive
`and Kidney Diseases convened a small panel of physicians and scientists working
`in the field to identify key unknowns and define possible “next steps” in advancing
`understanding of TSC- and mTOR-dependent renal phenotypes. TSC-associated
`renal disease, which affects ⬎85% of TSC patients, and was a major topic of
`discussion, focused on angiomyolipomas and epithelial cysts. The third major topic
`was the role of mTOR and mTOR inhibition in the pathogenesis and therapy of
`chronic renal disease. Renal cell carcinoma, while recognized as a manifestation of
`TSC that occurs in a small fraction of patients, was not the primary focus of this
`workshop and thus was omitted from panel discussions and from this report.
`
`angiomyolipoma; cyst; mTOR; tuberous sclerosis complex
`
`Angiomyolipomas
`
`Background. ANGIOMYOLIPOMAS can arise sporadically or as
`part of the tuberous sclerosis complex (TSC). Sporadic angio-
`myolipomas can be associated with translocations involving
`the TFE3 transcription factor (1) or the TSC genes (5). The
`panel focused primarily on TSC-associated angiomyolipomas.
`Angiomyolipomas are benign mesenchymal tumors composed
`of fat, smooth muscle, and abnormal vascular elements, all of
`which are known to arise from a common precursor cell (13).
`TSC patients typically have multiple, bilateral angiomyolipo-
`mas that can result in renal insufficiency, and these lesions
`
`Address for reprint requests and other correspondence: E. P. Henske, Karp
`Bldg., 6th Floor, One Blackfan Circle, Boston, MA 02115 (e-mail: EHenske
`@Partners.org).
`
`http://www.ajprenal.org
`
`have abnormal vasculature that can form aneurysms that can
`spontaneously hemorrhage, sometimes with life-threatening
`consequences (2). Clinical trials have clearly demonstrated that
`angiomyolipomas shrink in response to inhibitors of mamma-
`lian target of rapamycin complex 1 (mTORC1) (3, 4, 6, 8) and
`that the majority of the tumors return to original volume when
`the treatment is discontinued (4). Thus most patients appear to
`require continuous therapy to suppress angiomyolipoma size.
`Since the rapalog sirolimus typically induces a cytostatic re-
`sponse, it is likely that the decrease in tumor volume does not
`correlate with extensive apoptosis. However, because angiomyo-
`lipomas are highly vascular, mTORC1 inhibition could induce an
`apoptotic cellular response via an antiangiogenic mechanism.
`The allosteric inhibitors (rapalogs) and catalytic inhibitors
`(that impact both mTORC1 and 2) are different, with the
`catalytic inhibitors being much more effective in inhibiting
`
`F279
`
`Breckenridge Exhibit 1110
`Henske
`Page 001
`
`
`
`Downloaded from
`
`http://ajprenal.physiology.org/
`
` by 10.220.33.4 on January 8, 2017
`
`Review
`
`F280
`
`TSC, mTOR, AND THE KIDNEY
`
`mTOR activities based on in vitro studies. Preclinical work
`found equivalent benefit of allosteric and catalytic mTOR
`inhibitors in a murine model of TSC (10), but no clinical
`studies of the mTOR catalytic inhibitors involving TSC pa-
`tients have been completed.
`Key unanswered questions. The panel considered key issues
`that remain unaddressed related to the natural history and
`pathogenesis of angiomyolipomas, the evaluation/optimization
`of current angiomyolipoma therapies, and the development of
`novel angiomyolipoma treatment
`strategies, which were
`deemed essential to future progress. These issues are summa-
`rized in Fig. 1.
`Highest priority translational research initiatives. Multiple
`translational approaches were discussed by the panel to address
`the most critical unanswered questions outlined in Fig. 1. The
`highest priority initiatives are the following: 1) genetic analy-
`ses of angiomyolipomas from individual patients, including
`both indolent and rapidly growing tumors and including both
`the solid and vascular components; 2) identification of the
`cell-of-origin of angiomyolipomas, which could facilitate the
`development of animal models of angiomyolipomas; and 3) de-
`velopment of additional cell culture models of angiomyolipomas
`with bi-allelic TSC2 inactivation to facilitate translational and
`preclinical therapeutic advances.
`
`Highest priority clinical research initiatives. In parallel,
`multiple clinical research approaches were discussed to address
`unmet needs in the clinical care of individuals with angiomyo-
`lipomas. From these discussions, the highest priorities are the
`following.
`1) Identification of biomarkers to quantitatively and sensi-
`tively measure angiomyolipoma size and composition. Current
`technology does not easily quantitate the percentage of fat or
`smooth muscle components so that imaging between time
`points or patients can be accurately compared. Both imaging
`and “liquid” (plasma or urinary) biomarkers could be pivotal in
`monitoring angiomyolipoma burden of disease and therapeutic
`response. Imaging modalities that measure the effects of
`mTORC1 activation on cellular metabolism are particularly
`attractive. Serum biomarkers such as VEGF-D, which is a
`diagnostic biomarker and an indicator of therapeutic response
`in lymphangioleiomyomatosis (LAM) (25), a disease that
`shares cellular and genetic features with angiomyolipomas
`(12), should be sought, and markers like collagen IV should be
`pursued (3). Sensitive and specific biomarkers would facilitate
`trials to determine optimal dosing and duration of therapy, and
`to test novel therapies that might result in durable responses,
`ultimately resulting in shorter treatment intervals with associ-
`ated cost savings and a decreased risk of adverse effects.
`
`Natural History and
`Pathogenesis of Angiomyolipomas
`
`Evaluation/Optimization of Current
`Angiomyolipoma Therapies
`
`Development of Novel
`Angiomyolipoma Treatment Strategies
`
`(cid:129) Can early intervention with Rapalogs
` prevent the development of
`
` angiomyolipomas?
`(cid:129) Can addition of other therapeutic agents to
` Rapalogs produce a more robust and
` durable therapeutic response?
`
`(cid:129) Can mTORC1 inhibitors be targeted
`
` directly to angiomyolipomas, thereby
` avoiding systemic toxicity and increasing
` the dose delivered?
`(cid:129) Will catalytic inhibitors of mTOR, which
` inhibit both mTORC1 and mTORC2, be
` more effective than Rapalogs for
`
` angiomyolipoma therapy?
`(cid:129) Does mTOR-dependent feedback to AKT
` and MEK impact the clinical response to
` Rapalogs?
`(cid:129) How does activation of autophagy impact
` the therapeutic response to Rapalogs?
`
`(cid:129) Are angiomyolipomas “locally metastatic,”
` such that a single lesion can seed other
` lesions within the kidney?
`
`(cid:129) Are there genomic or epigenetic
` differences between indolent and locally
` aggressive angiomyolipomas?
`(cid:129) Is there a developmental window during
` which angiomyolipomas initiate?
`(cid:129) What are the mechanisms of aneurysm
` formation in angiomyolipomas?
`
`(cid:129) Are there gender differences in
` angiomyolipoma incidence, size, rate of
` growth, timing of growth, or risk of
`
` bleeding?
`(cid:129) Does blood pressure correlate with the risk
` of hemorrhage from angiomyolipomas?
`(cid:129) Can angiomyolipoma cells be detected in
` the circulation, as has been already shown
` in LAM, providing a “liquid biopsy” allowing
` cellular features of angiomyolipomas to be
` monitored in “real time.”
`
`(cid:129) Why is there such a variable response to
` mTORC1 inhibition?
` Some angiomyolipomas shrink in size
` while others do not, even within the
`
` same kidney.
` Does this reflect the composition of the
` tumor (fat-containing versus fat-poor) or
` its vasculature?
`
`(cid:129) Why do some tumors regrow when
` treatment is discontinued while others
` do not?
` What are the kinetics of the response,
` and is the earliest response vascular or
` cellular?
`(cid:129) What is the mechanism of response?
` Does the decrease in angiomyolipoma size
` with mTORC1 inhibition reflect primarily a
` reduction in individual cell volume, since
` mTORC1 is a key regulator of cell size, or
` a decrease in cell number?
`(cid:129) What is the minimum dose and duration of
` mTOR inhibitor therapy required to induce
` and maintain a reduction in angio-
`
` myolipoma size?
`(cid:129) What are the long-term benefits and risks
` of mTORC1 inhibitor therapy for
`
` angiomyolipomas, including the risk of
` hemorrhage, the impact on the
`
` aneurysmal vessels associated with
`
` angiomyolipomas, and the potential
`
` immunosuppressive effects?
`
`Fig. 1. Unanswered angiomyolipoma issues.
`
`AJP-Renal Physiol • doi:10.1152/ajprenal.00525.2013 • www.ajprenal.org
`
`Breckenridge Exhibit 1110
`Henske
`Page 002
`
`
`
`Downloaded from
`
`http://ajprenal.physiology.org/
`
` by 10.220.33.4 on January 8, 2017
`
`TSC, mTOR, AND THE KIDNEY
`
`Review
`
`F281
`
`Biomarkers may also identify the subset of angiomyolipomas
`with a more aggressive clinical phenotype, prioritizing patients
`with these tumors for earlier therapeutic intervention.
`2) There is an additional need for biomarkers that predict the
`risk of hemorrhage, which is a potentially life-threatening
`complication of angiomyolipomas. MRI analysis of aneurysm
`size/complexity may help define imaging biomarkers. Work
`with vascular-related markers may be helpful, and understand-
`ing the biology of aneurysms in the context of TSC will likely
`prove to be a critical step (3, 16, 24).
`3) Genome-wide studies to identify factors that modify the
`risk of angiomyolipoma development, the risk of angiomyoli-
`poma severity, the risk of angiomyolipoma hemorrhage, and/or
`the response to mTORC1 inhibitor therapy should be per-
`formed.
`4) The immunosuppressive effects of mTORC1 inhibitors,
`when used alone in TSC and related diseases, are unclear. The
`risk of infectious complications appears to be low, based on
`available data, but needs to be clarified through longer term
`observational studies.
`5) Natural history studies to identify the earliest develop-
`ment of angiomyolipomas, to determine whether spurts of
`growth can be defined in childhood, puberty, or young adult-
`hood, whether there are gender differences in the timing of
`angiomyolipoma growth, and whether bleeding risk and/or
`growth are correlated with clinical parameters including blood
`pressure, will be pivotal to future prevention trials.
`6) Designing and conducting a placebo-controlled, early-
`intervention study of mTORC1 inhibition for angiomyolipo-
`mas to evaluate its effectiveness as preventative therapy.
`7) Conducting clinical trials to evaluate agents that could
`synergize with mTORC1 inhibitors to produce more effective
`treatment, e.g., Hsp90 inhibitors, autophagy inhibitors, and
`mTOR kinase inhibitors.
`8) Determining the potential benefit of low-dose long-term
`therapies on inhibiting angiomyolipoma development, e.g.,
`
`metformin, NSAIDs, and low-dose or intermittent rapalog
`therapy.
`
`Renal Cystic Disease in TSC
`
`Background. Renal cystic disease in TSC is common, af-
`fecting ⬃50% of patients, ranging in severity from a single
`cyst to multiple, bilateral cystic disease (7, 9, 17). Individuals
`with the contiguous gene syndrome who carry deletions of both
`TSC2 and the adjacent PKD1 gene (⬍5% of TSC patients) can
`even develop severe very early onset polycystic kidney disease
`(18). The mTORC1 pathway has been implicated in the patho-
`genesis of renal cystic disease in autosomal dominant polycys-
`tic kidney disease (ADPKD) (21). To date, the use of mTOR
`inhibitors in treatment of ADPKD has yielded equivocal results
`(4, 5). The timing of mTORC1 inhibition may be pivotal in the
`response of ADPKD-associated cysts to therapy (20, 23).
`Defects in the primary cilium have been observed in TSC-
`deficient cells, including increased ciliary length (11). Together
`with the prominent cystic disease, these findings suggest that
`TSC may be a ciliopathy, yet the role of cilia in the initiation
`of, progression of, and therapy for renal cystic disease in TSC
`is not yet established.
`Key unanswered questions. The panel considered key issues
`involving TSC renal cystic disease that remain unaddressed
`and are essential to future progress. These issues are summa-
`rized in Fig. 2.
`Highest priority translational research initiatives. Multiple
`translational approaches to TSC renal cystic disease were
`discussed by the panel, with the highest priority initiatives
`being the following: 1) comprehensive genetic analysis of a
`large cohort of TSC patients with the polycystic kidney phe-
`notype to define in greater detail the spectrum of mutations that
`cause this manifestation and determine whether mutation of the
`contiguous TSC2 and PKD1 genes account for all of these
`patients; 2) development of an animal model of the TSC2/
`
`(cid:129) The therapeutic response of TSC-associated renal cysts to Rapalogs is largely unknown because the angiomyolipoma trials were not
` designed to assess cystic disease. A particularly important consideration is whether mTORC1 inhibitor therapy can prevent chronic
` kidney disease in patients with the TSC2/PKD1 contiguous gene deletion syndrome.
`(cid:129) The origin, developmental timing and natural history of cyst initiation and progression in TSC are poorly understood. To what extent do
` cysts initiate pre- and post-natally, and what factors and mechanisms are involved in their expansion?
` Do cysts in TSC originate from different segments of the kidney at different developmental time points?
` How does cyst formation and progression relate to angiomyolipoma development?
`(cid:129) The optimal therapeutic endpoints for cystic response are not defined. How should proliferation vs. secretion be considered when
` considering the response of cysts to therapy?
`(cid:129) Factors that may promote cyst progression are not defined. Is hypertension in TSC correlated with cystic disease?
` Does renal injury contribute to cyst progression?
`(cid:129) What accounts for the severity of cystic disease in the TSC2/PKD1 contiguous gene syndrome?
` The impact of co-deletion of TSC2 and PKD1 on a single copy of chromosome 16 in humans seems strikingly different than mutational
` inactivation of each gene on different copies of chromosome 16 in mouse models. Can a mouse model be generated with loss of both
` genes on a single allele to address this?
`(cid:129) There is considerable phenotypic variation in the degree of cyst formation in TSC patients. What is the basis for this variability and is it
` possible to carry out genetic studies of patients at the extremes of the phenotypic distribution?
`(cid:129) What are the functional consequences of ciliary dysfunction in TSC kidneys, and how does this contribute to cystogenesis?
`
`Fig. 2. Unanswered renal cystic disease issues.
`
`AJP-Renal Physiol • doi:10.1152/ajprenal.00525.2013 • www.ajprenal.org
`
`Breckenridge Exhibit 1110
`Henske
`Page 003
`
`
`
`Downloaded from
`
`http://ajprenal.physiology.org/
`
` by 10.220.33.4 on January 8, 2017
`
`Review
`
`F282
`
`TSC, mTOR, AND THE KIDNEY
`
`PKD1 contiguous gene deletion syndrome; 3) definition of the
`developmental timing and nephron segment-of-origin of cysts
`in TSC; 4) further examination of the connections between
`ciliary function and cyst formation in TSC; and 5) identifica-
`tion of factors (hypertension, injury) that promote cystogenesis
`in animal models of TSC.
`Highest priority clinical research initiatives. Multiple clin-
`ical research approaches to TSC renal cystic disease were
`considered with the highest priorities being 1) design of clin-
`ical trials to specifically determine whether and how cysts in
`TSC respond to rapalog therapy, including individuals with the
`TSC2/PKD1 contiguous gene syndrome, as the therapeutic
`response of the cystic disease has not been evaluated as an end
`point
`in previous studies; and 2) identification of factors
`including hypertension and renal injury that may promote cyst
`progression in TSC.
`
`Intrinsic Renal Disease Related to mTORC1 Inhibition
`
`Background. Although rapamycin appears to be minimally
`nephrotoxic when used alone, most of the data from humans
`are from studies in which it was used in combination with
`cyclosporine. Rapamycin was not associated with a significant
`increase in proteinuria during the EXIST2 trial of the rapalog
`everolimus for angiomyolipomas (3). However, this trial was
`of relatively short duration and included just over 100 patients.
`Thus the long-term effects of rapalogs as single agents on the
`kidney are not entirely understood. Prolonged treatment with
`mTORC1 inhibitors reduces the total expression of mTOR, as
`well as the expression of rictor and thus mTORC2 formation
`(19). Podocyte expression of nephrin, transient receptor poten-
`tial cation channel 6, and the cytoskeletal adaptor protein Nck
`are significantly decreased following prolonged exposure to an
`mTORC1 inhibitor (22). Furthermore, mTORC1 inhibition
`reduces podocyte adhesion and motility. Together, these ef-
`fects may have a long-term impact on the glomerular and
`tubular structures and deserve attention.
`Key unanswered questions. The panel considered key issues
`involving intrinsic renal disease related to mTORC1 that re-
`main unaddressed and that are essential to be understood as
`treatment may be prolonged. Questions that were discussed
`included the following. Does prolonged rapalog therapy induce
`proteinuria and/or other glomerular or tubular effects in hu-
`mans? Are there differential effects of mTORC1 vs. mTORC2
`inhibition on the kidney that could be relevant to future clinical
`trials involving catalytic mTOR kinase inhibitors?
`Highest priority translational and clinical research initiatives.
`Given that mTORC1 inhibitor therapy will be used in both
`children and adults with TSC and that there are many un-
`knowns related to the long-term impact on the kidney, the
`panel concluded that renal function and proteinuria should be
`monitored in a standardized, prospective manner in individuals
`receiving long-term rapalog therapy.
`
`Conclusions
`
`In summary, there was consensus that areas of high priority
`related to the roles of mTOR in renal disease include the
`following.
`Preclinical models of angiomyolipomas and renal cystic
`disease. Priorities for which the panel had clear consensus
`included identifying the cell-of-origin of angiomyolipomas,
`
`developing mouse models of angiomyolipomas, and develop-
`ing of additional cell lines derived from angiomyolipomas. A
`mouse model that recapitulates the severe, early-onset cystic
`disease observed in the TSC2/PKD1 contiguous gene syn-
`drome is likewise required. Further
`investigation of
`the
`nephron segment-of-origin, the developmental timing of cystic
`disease in TSC, and the role of ciliary dysfunction in TSC-
`associated cystogenesis is critical to the development of tar-
`geted therapy for TSC-associated renal cystic disease. It is also
`important to develop cell culture models for study of TSC
`using induced pluripotent stem cells (iPSC) from patients.
`Biomarkers. The panel determined that future research fo-
`cused on developing and refining imaging techniques to mon-
`itor disease progression, evaluating responses to therapy, and
`providing valuable natural history data would address key
`unmet needs. Focus areas included modalities that would allow
`more precise monitoring of tumor size, imaging with novel
`PET tracers that would allow differentiation between fat and
`other elements within angiomyolipomas, and serum biomark-
`ers of disease burden and therapeutic response are crucial.
`Similarly,
`imaging and biochemical biomarkers that could
`prognosticate and monitor therapy for the renal cystic disease
`are critical.
`Future clinical trials. The panel recommended that future
`clinical trials should include optimizing rapalog therapy by
`defining the minimum dose required that maintains a maximum
`response. Furthermore, identifying agents that could be com-
`bined with rapalogs or that could be used individually to induce
`a more complete and/or durable response could prove to be
`pivotal in allowing periodic or one-time, rather than life-long,
`therapy. In addition, determining whether catalytic mTOR
`inhibitors induce a more complete and/or durable response
`compared with rapalogs is a high priority.
`Prevention. In general, current studies have focused on the
`treatment of large or enlarging angiomyolipomas. Future stud-
`ies should focus on the prevention of angiomyolipomas, which
`would require additional natural history information, including
`defining whether there is a “window” of more rapid growth of
`angiomyolipomas during development. Although there is an
`anecdotal sense that angiomyolipomas can grow more rapidly
`during adolescence and early adulthood, this is not well de-
`fined. Epithelial cysts in TSC remain understudied, and the
`effects of rapalogs on cyst progression are unknown. Individ-
`uals with the TSC2/PKD1 contiguous gene syndrome, who are
`at risk for early-onset, severe polycystic kidney disease, rep-
`resent a population in whom prevention studies are a high
`priority.
`
`ACKNOWLEDGMENTS
`
`This work was submitted on behalf of the National Institute of Diabetes and
`Digestive and Kidney Diseases (NIDDK) TSC Working Group: E. Henske,
`Brigham and Women’s Hospital; J. Bissler, Le Bonheur Children’s Hospital;
`R. Rasooly, NIDDK; Chris Kingswood, Royal Sussex County Hospital,
`Brighton, UK; Elizabeth Thiele, Massachusetts General Hospital; Julian Samp-
`son, Institute of Medical Genetics, Cardiff University, Cardiff, UK; David
`Kwiatkowski, Harvard Medical School; Brendan Manning, Harvard School of
`Public Health; B. Siroky, Cincinnati Children’s Hospital Medical Center;
`Thomas Weimbs, University of California Santa Barbara; and Leon Murphy,
`Novartis Pharmaceuticals.
`
`DISCLOSURES
`
`No conflicts of interest, financial or otherwise, are declared by the authors.
`
`AJP-Renal Physiol • doi:10.1152/ajprenal.00525.2013 • www.ajprenal.org
`
`Breckenridge Exhibit 1110
`Henske
`Page 004
`
`
`
`Downloaded from
`
`http://ajprenal.physiology.org/
`
` by 10.220.33.4 on January 8, 2017
`
`TSC, mTOR, AND THE KIDNEY
`
`Review
`
`F283
`
`AUTHOR CONTRIBUTIONS
`
`Author contributions: E.P.H., B.J.S., and J.J.B. drafted manuscript; E.P.H.,
`R.R., B.J.S., and J.J.B. edited and revised manuscript; E.P.H., R.R., B.J.S., and
`J.J.B. approved final version of manuscript.
`
`REFERENCES
`
`1. Argani P, Aulmann S, Illei PB, Netto GJ, Ro J, Cho HY, Dogan S,
`Ladanyi M, Martignoni G, Goldblum JR, Weiss SW. A distinctive
`subset of PEComas harbors TFE3 gene fusions. Am J Surg Pathol 34:
`1395–1406, 2010.
`2. Bissler JJ, Kingswood JC. Renal angiomyolipomata. Kidney Int 66:
`924 –934, 2004.
`3. Bissler JJ, Kingswood JC, Radzikowska E, Zonnenberg BA, Frost M,
`Belousova E, Sauter M, Nonomura N, Brakemeier S, de Vries PJ,
`Whittemore VH, Chen D, Sahmoud T, Shah G, Lincy J, Lebwohl D,
`Budde K. Everolimus for angiomyolipoma associated with tuberous
`sclerosis complex or sporadic lymphangioleiomyomatosis (EXIST-2): a
`multicentre, randomised, double-blind, placebo controlled trial. Lancet
`381: 817–824, 2013.
`4. Bissler JJ, McCormack FX, Young LR, Elwing JM, Chuck G, Leon-
`ard JM, Schmithorst VJ, Laor T, Brody AS, Bean J, Salisbury S,
`Franz DN. Sirolimus for angiomyolipoma in tuberous sclerosis complex
`or lymphangioleiomyomatosis. N Engl J Med 358: 140 –151, 2008.
`5. Carsillo T, Astrinidis A, Henske EP. Mutations in the tuberous sclerosis
`complex gene TSC2 are a cause of sporadic pulmonary lymphangioleio-
`myomatosis. Proc Natl Acad Sci USA 97: 6085–6090, 2000.
`6. Dabora SL, Franz DN, Ashwal S, Sagalowsky A, DiMario FJ Jr, Miles
`D, Cutler D, Krueger D, Uppot RN, Rabenou R, Camposano S, Paolini
`J, Fennessy F, Lee N, Woodrum C, Manola J, Garber J, Thiele EA.
`Multicenter phase 2 trial of sirolimus for tuberous sclerosis: kidney
`angiomyolipomas and other tumors regress and VEGF- D levels decrease.
`PloS One 6: e23379, 2011.
`7. Dabora SL, Jozwiak S, Franz DN, Roberts PS, Nieto A, Chung J,
`Choy YS, Reeve MP, Thiele E, Egelhoff JC, Kasprzyk-Obara J,
`Domanska-Pakiela D, Kwiatkowski DJ. Mutational analysis in a cohort
`of 224 tuberous sclerosis patients indicates increased severity of TSC2,
`compared with TSC1, disease in multiple organs. Am J Hum Genet 68:
`64 –80, 2001.
`8. Davies DM, Johnson SR, Tattersfield AE, Kingswood JC, Cox JA,
`McCartney DL, Doyle T, Elmslie F, Saggar A, de Vries PJ, Sampson
`JR. Sirolimus therapy in tuberous sclerosis or sporadic lymphangioleio-
`myomatosis. N Engl J Med 358: 200 –203, 2008.
`9. Ewalt DH, Sheffield E, Sparagana SP, Delgado MR, Roach ES. Renal
`lesion growth in children with tuberous sclerosis complex. J Urol 160:
`141–145, 1998.
`10. Guo Y, Kwiatkowski DJ. Equivalent benefit of rapamycin and a potent
`mTOR ATP-competitive inhibitor, MLN0128 (INK128),
`in a mouse
`model of tuberous sclerosis. Mol Cancer Res 11: 467–473, 2013.
`11. Hartman TR, Liu D, Zilfou JT, Robb V, Morrison T, Watnick T,
`Henske EP. The tuberous sclerosis proteins regulate formation of the
`primary cilium via a rapamycin-insensitive and polycystin 1-independent
`pathway. Hum Mol Genet 18: 151–163, 2009.
`12. Henske EP, McCormack FX. Lymphangioleiomyomatosis—a wolf in
`sheep’s clothing. J Clin Invest 122: 3807–3816, 2012.
`
`13. Karbowniczek M, Yu J, Henske EP. Renal angiomyolipomas from
`patients with sporadic lymphangiomyomatosis contain both neoplastic and
`non-neoplastic vascular structures. Am J Pathol 162: 491–500, 2003.
`14. Krueger DA, Care MM, Holland K, Agricola K, Tudor C, Mangesh-
`kar P, Wilson KA, Byars A, Sahmoud T, Franz DN. Everolimus for
`subependymal giant-cell astrocytomas in tuberous sclerosis. N Engl J Med
`363: 1801–1811, 2010.
`15. McCormack FX, Inoue Y, Moss J, Singer LG, Strange C, Nakata K,
`Barker AF, Chapman JT, Brantly ML, Stocks JM, Brown KK, Lynch
`JP, 3rd Goldberg HJ, Young LR, Kinder BW, Downey GP, Sullivan
`EJ, Colby TV, McKay RT, Cohen MM, Korbee L, Taveira-DaSilva
`AM, Lee HS, Krischer JP, Trapnell BC. Efficacy and safety of sirolimus
`in lymphangioleiomyomatosis. N Engl J Med 364: 1595–1606, 2011.
`16. O’Brien K, Leach J, Jones B, Bissler J, Zuccarello M, Abruzzo T.
`Calcifications associated with pediatric intracranial arterial aneurysms:
`incidence and correlation with pathogenetic subtypes. Child’s Nerv Syst
`29: 643–649, 2013.
`17. Rakowski SK, Winterkorn EB, Paul E, Steele DJ, Halpern EF, Thiele
`EA. Renal manifestations of tuberous sclerosis complex: Incidence, prog-
`nosis, and predictive factors. Kidney Int 70: 1777–1782, 2006.
`18. Sampson JR, Maheshwar MM, Aspinwall R, Thompson P, Cheadle
`JP, Ravine D, Roy S, Haan E, Bernstein J, Harris PC. Renal cystic
`disease in tuberous sclerosis: role of the polycystic kidney disease 1 gene.
`Am J Hum Genet 61: 843–851, 1997.
`19. Sarbassov DD, Ali SM, Sengupta S, Sheen JH, Hsu PP, Bagley AF,
`Markhard AL, Sabatini DM. Prolonged rapamycin treatment inhibits
`mTORC2 assembly and Akt/PKB. Mol Cell 22: 159 –168, 2006.
`20. Serra AL, Poster D, Kistler AD, Krauer F, Raina S, Young J, Rentsch
`KM, Spanaus KS, Senn O, Kristanto P, Scheffel H, Weishaupt D,
`Wuthrich RP. Sirolimus and kidney growth in autosomal dominant
`polycystic kidney disease. N Engl J Med 363: 820 –829.
`21. Shillingford JM, Murcia NS, Larson CH, Low SH, Hedgepeth R,
`Brown N, Flask CA, Novick AC, Goldfarb DA, Kramer-Zucker A,
`Walz G, Piontek KB, Germino GG, Weimbs T. The mTOR pathway is
`regulated by polycystin-1, and its inhibition reverses renal cystogenesis in
`polycystic kidney disease. Proc Natl Acad Sci USA 103: 5466 –5471,
`2006.
`22. Vollenbroker B, George B, Wolfgart M, Saleem MA, Pavenstadt H,
`Weide T. mTOR regulates expression of slit diaphragm proteins and
`cytoskeleton structure in podocytes. Am J Physiol Renal Physiol 296:
`F418 –F426, 2009.
`23. Walz G, Budde K, Mannaa M, Nurnberger J, Wanner C, Sommerer
`C, Kunzendorf U, Banas B, Horl WH, Obermuller N, Arns W,
`Pavenstadt H, Gaedeke J, Buchert M, May C, Gschaidmeier H,
`Kramer S, Eckardt KU. Everolimus in patients with autosomal dominant
`polycystic kidney disease. N Engl J Med 363: 830 –840.
`24. Williams JM, Racadio JM, Johnson ND, Donnelly LF, Bissler JJ.
`Embolization of renal angiomyolipomata in patients with tuberous scle-
`rosis complex. Am J Kidney Dis 47: 95–102, 2006.
`25. Young LR, Vandyke R, Gulleman PM, Inoue Y, Brown KK, Schmidt
`LS, Linehan WM, Hajjar F, Kinder BW, Trapnell BC, Bissler JJ,
`Franz DN, McCormack FX. Serum vascular endothelial growth factor-D
`prospectively distinguishes lymphangioleiomyomatosis from other dis-
`eases. Chest 138: 674 –681, 2010.
`
`AJP-Renal Physiol • doi:10.1152/ajprenal.00525.2013 • www.ajprenal.org
`
`Breckenridge Exhibit 1110
`Henske
`Page 005
`
`